Smoothelin-like 1 knockout mice display sex-dependent alterations in blood flow and cardiac function.

Smoothelin-like 1 (SMTNL1) modulates the contractile performance of smooth muscle and thus has a key role in vascular homeostasis. Elevated vascular tone, recognized as a contributor to the development of progressive cardiac dysfunction, was previously found with SMTNL1 deletion. In this study, we assessed cardiac morphology and function of male and female, wild-type (Smtnl1+/+) and global SMTNL1 knockout (Smtnl1-/-) mice at 10 weeks of age. Gross dissection revealed distinct cardiac morphology only in males; Smtnl1-/- hearts were significantly smaller than Smtnl1+/+, but the left ventricle (LV) proportion of heart mass was greater. Male Smtnl1-/- mice also displayed increased ejection fraction and fractional shortening, as well as elevated aortic and pulmonary flow velocities. The impact of cardiac stress with pressure overload by transverse aortic constriction (TAC) was examined in male mice. With TAC banding, systolic function was preserved, but the LV filling pressure was selectively elevated due to relaxation impairment. Smtnl1-/- mice displayed higher early/passive filling velocity of LV/early mitral annulus velocity ratio (E/E' ratio) and myocardial performance index along with a prolonged isovolumetric relaxation time. Taken together, the findings support a novel, sex-dimorphic role for SMTNL1 in modulating cardiac structure and function of mice.

An immunohistochemical study of diffuse large B-cell lymphoma with molecular subtyping based on Hans algorithm.

INTRODUCTION: Diffuse large B-cell lymphoma (DLBCL) is the most common type of B-cell lymphoma accounting for about 40% of all lymphomas. The international prognostic index (IPI), which relies on clinical and laboratory parameters, is used as a prognostic tool in DLBCL. In this study, we have included cases of DLBCL not otherwise specified (NOS) type to test the usefulness of the biological subclassification of DLBCL by immunohistochemistry (IHC) using the Hans algorithm into the germinal center B-like (GCB) type and nongerminal center (non-GCB) type. We correlated the subtypes with the demographics, site of involvement, IPI scores, and stage. OBJECTIVES: 1. To study the immunohistochemical profile of DLBCL. 2. To classify patients with DLBCL into germinal center and nongerminal center subtypes. 3. To review the clinical presentations, clinical staging, and follow-up data in cases of DLBCL. Materials and Methods: A total of 152 cases of DLBCL reported from January 2010 to March 2018 were included in this study. Clinical data, treatment details, and follow-up were reviewed. Apart from the routine IHC markers for DLBCL, additional markers CD10, BCL6, and MUM 1 were performed to classify DLBCL into GCB type and non-GCB type using the Hans algorithm. RESULTS: The median age of presentation was 53 years with male-to-female ratio of 2:1. Most of the patients presented with nodal involvement (56.6%); cervical lymph node is the most common site (46.5%). The majority of the patients presented in Ann Arbor stage 1 (44.8%). According to the international prognostic index, 34.8% had a score of 3 (high intermediate). After IHC studies, GCB subtype (51.5%) of DLBCL emerged as more common than non-GCB type (48.5%). The two subtypes differed significantly with regard to sex (i.e. GCB type was more common in males and non-GCB type in females) and showed no significance with regard to any of the other clinical features and prognostic parameters evaluated. CONCLUSION: Our study showed that IHC was a useful tool to subclassify DLBCL into GCB and non-GCB subtypes and may be easily incorporated in routine clinical practice.

Endosulfan causes oxidative stress in the liver and brain that involves inhibition of NADH dehydrogenase and altered antioxidant enzyme status in rat.

Endosulfan, a neurotoxic, highly persistent organochlorine insecticide, is known for its acute and chronic toxicity. We have shown that a single sublethal dose of endosulfan caused high induction of oxidative stress in the liver and brain by altering the antioxidant status, as shown by reduction in the antioxidant enzymes SOD, GPx, GST, GR along with increased ROS and lipid peroxidation. The cerebral region in the brain showed a higher level of oxidative stress than the cerebellum, revealing differential sensitivity of the brain regions to endosulfan. Depletion of natural antioxidants causes the imbalance of redox status in cells, and the role of mitochondrial distress causally related to the cellular oxidative stress in vivo is not well understood. We have shown that reduction in the mitochondrial NADH dehydrogenase activity in the brain is associated with the induction of ROS in endosulfan-treated rats. Although oxidative stress is induced in both the liver and brain, the oxidative damage to the brain has implications for the toxic outcome in view of the brain's lower antioxidant defenses and high oxygen consumption.

Thrombin-Mediated Formation of Globular Adiponectin Promotes an Increase in Adipose Tissue Mass.

A decrease in the circulating levels of adiponectin in obesity increases the risk of metabolic complications, but the role of globular adiponectin, a truncated form produced by proteolytic cleavage, has not been defined. The objective of this investigation was to determine how globular adiponectin is generated and to determine whether this process impacts obesity. The cleavage of recombinant full-length adiponectin into globular adiponectin by plasma in vitro was used to identify Gly-93 as the N-terminal residue after proteolytic processing. The amino acid sequence of the cleavage site suggested thrombin was the protease responsible for cleavage, and inhibitors confirmed its likely involvement. The proteolytic site was modified, and this thrombin-resistant mutant protein was infused for 4 weeks into obese adiponectin-knockout mice that had been on a high-fat diet for 8 weeks. The mutation of the cleavage site ensured that globular adiponectin was not generated, and thus did not confound the actions of the full-length adiponectin. Mice infused with the mutant adiponectin accumulated less fat and had smaller adipocytes compared to mice treated with globular adiponectin, and concurrently had elevated fasting glucose. The data demonstrate that generation of globular adiponectin through the action of thrombin increases both adipose tissue mass and adipocyte size, but it has no effect on fasting glucose levels in the context of obesity.

Oxidative stress-mediated cytotoxicity of Endosulfan is causally linked to the inhibition of NADH dehydrogenase and Na+, K+-ATPase in Ehrlich ascites tumor cells.

Oxidative stress in cells caused by excessive production of reactive oxygen species (ROS) and decreased antioxidant defense is implicated in the cytotoxicity of xenobiotics including drugs and environmental chemicals. Endosulfan, a highly toxic organochlorine insecticide, causes cytotoxic cell death by inducing oxidative stress. We have investigated the biochemical basis of induction of oxidative stress, involving the role of NADH dehydrogenase and the possible role of Na+, K+-ATPase in endosulfan cytotoxicity and, whether the cytotoxicity could be attenuated by targeting ROS induction using the natural flavonoid antioxidant, quercetin, in Ehrlich ascites tumor (EAT) cells. Exposure of cells to endosulfan caused cytotoxic cell death (necrosis) which was associated with induction of ROS, lipid peroxidation as well as a reduction in glutathione levels, concomitant with loss of NADH dehydrogenase and Na+, K+-ATPase activity in a dose-dependent manner, indicating that oxidative stress and perturbation of membrane function are the major causes of endosulfan cytotoxicity. Our results showed that quercetin, protected against endosulfan-induced cytotoxicity and significantly abrogated oxidative stress, and ameliorated the inhibition of NADH dehydrogenase and Na+, K+-ATPase activity in EAT cells. Our study presents evidence that NADH dehydrogenase inhibition plays an important role in oxidative stress-mediated cytotoxicity, and perturbed membrane function as evident from inhibition of sodium-potassium pump is involved in cytotoxic cell death.

Smoothelins and the Control of Muscle Contractility.

Smooth muscle cells display distinctive expression and organization of contractile filament proteins, which reflect a unique method of contractile regulation. As the focus of this review, the smoothelin and smoothelin-like family members represent a family of poorly understood muscle proteins that appear to act as structural components of the contractile apparatus. The protein family is characterized by the presence a single C-terminal type-2 calponin homology (CH) domain. Often used as the preferred marker of differentiated contractile smooth muscle cells, smoothelin A and B (SMTN-A and SMTN-B) may influence the contractile potential of smooth muscle cells. The more recently identified smoothelin-like proteins (SMTNL1 and SMTNL2) have more diverse functional implications. SMTNL1 is linked to the regulation of smooth muscle contractility and adaptations of both smooth and skeletal muscle to hypertension, pregnancy, and exercise training. The SMTNL1 protein is suggested to play multiple roles in muscle through functional interactions with contractile regulators (e.g., calmodulin, tropomyosin, and myosin phosphatase) as well as transcriptional control of the contractile phenotype and Ca2+-sensitizing capacity. These effects are associated with acute, reversible changes to the contractile state or long-term adaptations mediated by transcriptional changes in expression of contractile proteins. SMTNL2 remains essentially uncharacterized; however, its expression is high in skeletal muscle and could be associated with differentiating myocytes. Finally, emerging opportunities exist to understand the significance of smoothelins as disease-associated markers and in some cases as specific modulators of pathophysiology.

STAT3 and MCL-1 associate to cause a mesenchymal epithelial transition.

Embryo implantation is effected by a myriad of signaling cascades acting on the embryo-endometrium axis. Here we show, by using MALDI TOF analysis, far-western analysis and colocalization and co-transfection studies, that STAT3 and MCL-1 are interacting partners during embryo implantation. We show in vitro that the interaction between the two endogenous proteins is strongly regulated by estrogen and progesterone. Implantation, pregnancy and embryogenesis are distinct from any other process in the body, with extensive, but controlled, proliferation, cell migration, apoptosis, cell invasion and differentiation. Cellular plasticity is vital during the early stages of development for morphogenesis and organ homeostasis, effecting the epithelial to mesenchymal transition (EMT) and, the reverse process, mesenchymal to epithelial transition (MET). STAT3 functionally associates with MCL-1 in the mammalian breast cancer cell line MCF7 that overexpresses STAT3 and MCL-1, which leads to an increased rate of apoptosis and decreased cellular invasion, disrupting the EMT. Association of MCL-1 with STAT3 modulates the normal, anti-apoptotic, activity of MCL-1, resulting in pro-apoptotic effects. Studying the impact of the association of STAT3 with MCL-1 on MET could lead to an enhanced understanding of pregnancy and infertility, and also metastatic tumors.

CrkL is a co-activator of estrogen receptor alpha that enhances tumorigenic potential in cancer.

Signaling via estrogen receptor (ER) occurs by interacting with many proteins. Nuclear interactome analysis of ERα in an embryo implantation model revealed the association of chicken tumor virus no. 10 regulator of kinase like (CrkL) with ERα, which was further validated by mammalian two-hybrid assay as well as coimmunoprecipitation and colocalization. Mutation in LPALL motif of CrkL disrupts the ERα-CrkL interaction and its transactivation potential, thereby suggesting that the interaction is mediated via its single ER binding motif, Leu-Pro-Ala-Leu-Leu (LXXLL) motif in the sarcoma homology (SH)2 domain. CrkL deletion constructs of SH2 domain target to the nucleus due to presence of nuclear localization signal. Interestingly, the SH2-SH3 (N terminal) construct shows an increased transactivation potential like CrkI. Weak interaction capability of mutated ERα-Y538F with CrkL validates that CrkL interacts with ERα via its YDLL motif at Tyr 541. In an attempt to understand the physiological relevance of this association, we investigated the impact on cell proliferation using a cancer model, because events associated in the process of pregnancy and cancer are analogous. Also, overexpression of CrkL is frequently associated with tumorigenesis. However, its significance in hormone-regulated cancers still remains obscure. Here, we demonstrate that association of ERα and CrkL directly enhances the tumorigenic potential of CrkL, thus pointing to its role in cell proliferation. In human endometrial cancers, we observed a strong association between ERα and CrkL levels. Thus, the molecular signaling set off by ERα and CrkL association may have a central role in pregnancy and cancer, two events which share parallels in growth, invasion, and immune tolerance.